June 2023
Volume 64, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2023
Differential gene expression patterns between cultured corneal keratocytes, fibroblasts, and myofibroblasts
Author Affiliations & Notes
  • Kara Poole
    Ophthalmology, The University of Texas Southwestern Medical Center, Dallas, Texas, United States
  • Krithika Iyer
    Biomedical Engineering, The University of Texas at Dallas, Richardson, Texas, United States
    Ophthalmology, The University of Texas Southwestern Medical Center, Dallas, Texas, United States
  • Steffi Daniel
    Ophthalmology, The University of Texas Southwestern Medical Center, Dallas, Texas, United States
  • Victor Varner
    Biomedical Engineering, The University of Texas at Dallas, Richardson, Texas, United States
    Surgery, The University of Texas Southwestern Medical Center, Dallas, Texas, United States
  • David Schmidtke
    Biomedical Engineering, The University of Texas at Dallas, Richardson, Texas, United States
    Surgery, The University of Texas Southwestern Medical Center, Dallas, Texas, United States
  • Matthew Petroll
    Ophthalmology, The University of Texas Southwestern Medical Center, Dallas, Texas, United States
    Biomedical Engineering, The University of Texas Southwestern Medical Center, Dallas, Texas, United States
  • Footnotes
    Commercial Relationships   Kara Poole None; Krithika Iyer None; Steffi Daniel None; Victor Varner None; David Schmidtke None; Matthew Petroll None
  • Footnotes
    Support  NIH grants R01 EY030190, R01 EY013322, P30 EY030413
Investigative Ophthalmology & Visual Science June 2023, Vol.64, 4595. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Kara Poole, Krithika Iyer, Steffi Daniel, Victor Varner, David Schmidtke, Matthew Petroll; Differential gene expression patterns between cultured corneal keratocytes, fibroblasts, and myofibroblasts. Invest. Ophthalmol. Vis. Sci. 2023;64(8):4595.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

Purpose : After stromal injury, the release of pro-inflammatory cytokines and growth factors typically results in activation of quiescent keratocytes to migratory fibroblast and/or fibrotic myofibroblast phenotypes. To our knowledge, there has not been a comprehensive analysis of the changes in gene expression that underlie these distinct wound healing phenotypes. The goal of this study is to identify transcriptional differences and curate characteristic gene expression profiles for cultured corneal keratocytes, fibroblasts, and myofibroblasts.

Methods : Primary rabbit corneal keratocytes were cultured for 5 days in serum-free media (SF), serum-containing media (S+), or SF with TGF-β1, to produce keratocytes, fibroblasts, and myofibroblasts, respectively. Four samples of total RNA were collected for each treatment group using the BioRad Aurum Total RNA Mini Kit, and two experimental replicates were sent to Novogene for bulk RNA-sequencing. Subsequent bioinformatics analysis included gene expression quantification, differential expression (DE), and functional analysis.

Results : Overall, there were 7693 significant (Padj<0.05 and log2(FoldChange)>1) DE genes between keratocytes and fibroblasts, 4430 between keratocytes and myofibroblasts, and 5704 between fibroblasts and myofibroblasts. When comparing S+ and TGF-β1 conditions to SF (control), genes characteristic of a quiescent keratocyte phenotype were generally downregulated (e.g. KERA and ALDH1A1). In S+ conditions, initial analysis revealed upregulation of genes associated with DNA replication, metabolism, and ECM-receptor interactions, and downregulation of cell-cell adhesion markers. There was also DE of genes associated with PI3K-Akt and Rap1 signaling. In TGF-β1 conditions, we observed upregulation of genes associated with focal adhesion formation and ECM remodeling. There was also DE of genes associated with PI3K-Akt, JAK-STAT, MAPK, PPAR, Rap1, and TNF signaling pathways. Additionally, for genes related to growth factor activity, there was an upregulation of TGF-β and PDGF family genes and a downregulation of FGF genes.

Conclusions : These data demonstrate that there are distinct gene expression profiles for cultured keratocytes, fibroblasts, and myofibroblasts. Further analysis of these transcriptomes may lead to identification of new gene markers/targets and discovery of novel signaling pathways associated with these phenotypes.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×